Generation and preliminary application of rabbit antihuman polyclonal antibodies against DR5

Objective To generate and identify the rabbit polyclonal antibodies against sDR5. Methods Rabbits were immunized with sDR5 for 3 times. The polyclonal antibodies from the collected antiserum were purified with protein G. Specialty of the polyclonal antibodies was analyzed by indirect-ELISA; apoptotic effects of the polyclonal antibodies on Jurkat cells were detected by FCM analysis; Bcl 2,CytC,caspase3,Bax,and caspase9 were detected with Western blotting. Results The titers of purified polyclonal antibodies were 1∶32 000. Western blot results showed that the antibodies could recognize the protein with molecular weight of 26 000 (sDR5). Apoptosis of Jurkat cells treated by the polyclonal antibodies was increased obviously. Specificity of Bcl 2,CytC,caspase3,Bax,and caspase9 were detected,and they changed with the time of polyclonal antibodies treatment. Conclusion Polyclonal antibodies against sDR5 has been prepared successfully. The polyclonal antibodies may be a useful tool in investigating tumor's therapy by using DR5 as target molecule and exploring the functional domain of DR5.